POSTER: High Throughput Proteomic Applications Using anti-DYKDDDDK Magnetic Agarose

C.E. Credits: P.A.C.E. CE Florida CE
Speaker
  • R&D Staff Scientist, Protein and Cell Analysis, Thermo Fisher Scientific
    Biography
      Betsy Benton completed her PhD and postdoctoral research at the University of Wisconsin-Madison in the fields of biochemistry and human oncology. She joined Thermo Fisher Scientific in 2000, where she has developed numerous products in the areas of protein purification (with both magnetic beads and resin supports), protein sample preparation, and protein detection.

    Abstract

    Affinity purification, immunoprecipitation, and mass spectrometry analysis are essential tools used in proteomic studies. A successful outcome is facilitated by high-capacity supports, efficient sample preparation, and reproducibility provided by automation. We demonstrate that an anti-DYKDDDDK magnetic agarose fulfills these criteria. The DYKDDDDK epitope tag is commonly used in protein expression systems as a short purification handle. Using an antibody against a DYKDDDDK tag covalently attached to high-binding capacity magnetic agarose, we demonstrate effective purification of recombinant proteins expressed in bacteria, the Invitrogen™ ExpiCHO™ Expression System, and by a HeLa in vitro translation (cell-free) system. Biological activity was demonstrated for ExpiCHO-expressed DYKDDDDK–Tumor Necrosis Factor alpha (TNFα) which was used to successfully stimulate cells and activate Nuclear Factor kappa B (NFkB) p65. In addition, successful co-immunoprecipitation of DYKDDDDK-tagged BAD with 14-3-3 proteins was confirmed using an optimized sample preparation protocol in conjunction with the Thermo Scientific™ KingFisher™ Flex Magnetic Particle Processor and mass spectrometry. In summary, anti-DYKDDDDK magnetic agarose can be used to effectively isolate, purify, and enrich functional protein targets in a variety of proteomic applications.

    Learning Objectives:

    • Learn how to automate protein applications using cell-based and cell-free systems with magnetic beads

    Show Resources
    You May Also Like
    JAN 23, 2020 9:00 AM PST
    C.E. CREDITS
    JAN 23, 2020 9:00 AM PST
    DATE: January 23, 2020 TIME: 9:00am PST, 12:00pm EST...
    APR 07, 2020 8:00 AM PDT
    C.E. CREDITS
    APR 07, 2020 8:00 AM PDT
    DATE: April 7, 2020 TIME: 8:00am PT, 11:00am ET This webinar sets out to establish why quality control is key to robust, reliable, reproducible science. We will look at best practice criteri...
    MAY 08, 2020 10:00 AM PDT
    C.E. CREDITS
    MAY 08, 2020 10:00 AM PDT
    DATE: May 8, 2020 TIME: 10:00am PT, 11:00am MT, 1:00pm ET The application of next generation sequencing to interrogate immune repertoires and methods in which these highly complex dataset...
    MAY 12, 2020 9:00 AM PDT
    C.E. CREDITS
    MAY 12, 2020 9:00 AM PDT
    NEW DATE: Date: May 12, 2020 Time: 9:00am PDT, 12:00pm EDT...
    APR 20, 2020 8:00 AM PDT
    C.E. CREDITS
    APR 20, 2020 8:00 AM PDT
    This talk will cover different approaches to emergency hemostasis testing for patients that are actively bleeding due to trauma, surgery, obstetrical emergencies and other causes. The talk i...
    JUN 23, 2020 10:00 AM PDT
    Add to Calendar Select one of the following: iCal Google Calendar Outlook Calendar Yahoo Calendar
    C.E. CREDITS
    JUN 23, 2020 10:00 AM PDT
    Add to Calendar Select one of the following: iCal Google Calendar Outlook Calendar Yahoo Calendar
    DATE: June 23, 2020 TIME: 10:00am PT Human mesenchymal stromal or stem cells (MSCs)-based immunomodulation treatment has been proposed as a suitable therapeutic approach for many diseases, s...
    Loading Comments...
    Show Resources